1. Field of the Invention
The present invention relates to solutions for detecting cellular material, and involves imaging techniques, image processing, optics, fluorescence and biochemistry.
2. Background Art
Micro-arrays are matrices of bio molecules spots deposited on a substrate that usually has the appearance of a microscope slide.
Micro-array technology was initially developed for genomics studies in order to realize massively parallel genetic assays for research applications such as human genome sequencing. DNA micro-array is a collection of microscopic DNA spots attached to a solid surface, such as glass, plastic or silicon chip forming an array for the purpose of expression profiling, monitoring expression levels for thousands of genes simultaneously. In one such application, molecules which are immobilized are single-stranded DNA molecules. A solution of probe molecules, which are fluorescently labeled single stranded DNA, is then made to interact with the micro-array. The probed DNA will then specifically bind to spots that are formed of complementary strands therefore revealing that those spots have a sequence that is similar to the probe. The arrays are then dried and read by a fluorescence detection system.
DNA micro-array fluorescent signals can be very weak which has led micro-array scanners to evolve into sophisticated and expensive confocal laser scanning systems for optimal sensibility, or cooled CCD camera systems for fast readout. These types of systems produce an image of the whole micro-array with a resolution of a few hundred pixels per spot. The imaging system's capability is used for compensating for variations in locations of the spots within the micro-array disposition, and for segmenting the spots from their surrounding background signal.
Recently, proteins have been immobilized on substrates to produce protein arrays. The deposited proteins can be antibodies targeted to specifically bind to viruses, toxins and micro-organism. The assay consists in letting a solution of unknown biological agent interact and specifically bind with the proteins on the array. If the unknown agents were previously fluorescently labeled, the micro-array can be analyzed right away, otherwise the array is revealed with a solution of fluorescently labeled antibodies that are also specific to the targeted biological agent. Antibodies or other types of capture molecules which are more similar in their nature, such as peptides or aptamers, can also be used to target agents.
Conventional micro-array scanners designed for DNA micro-arrays as detailed above, are still used for the analysis of protein based micro-arrays even though the requirements for these new applications are very different. The signal read by a conventional scanner is a measure of the total signal emanating from one spot, and can thus be affected by spurious contributions. Impurities, such as dusts and biological residues from the host solution, generally fluoresce and therefore contribute to the measured intensity of the spot's signal, in a way that is not related to the targeted agent. Furthermore, the efficiency of the fluorescence labeling of the revealing antibodies can vary greatly from experiment to experiment, leading to added variability which impairs the limit of detection.